HIV-1 gp41 contains highly conserved regions that are attractive targets for vaccine design. One such region is the membrane-proximal external region (MPER) of gp41, and the human monoclonal antibodies (mAbs) that bind to it, 2F5, 4E10 and Z13, neutralize primary isolates of HIV-1 from different clades. In addition, the N-heptad repeat (NHR) region of gp41 is a target of fusion inhibitor drugs and neutralizing mAbs. (In our preliminary studies, we have identified an affinity-improved version of Z13 (Z13e1), as well as rabbit mAbs and a human HIV-1 neutralizing mAb that were elicited against the NHR region of gp41.) Eliciting broadly neutralizing Abs against gp41 has been difficult due in part to heterogeneity in gp41 structure and a poor understanding of the specific structure and presentation of the neutralizing epitopes on HIV-1 gp41. We have designed a series of gp41 mimetics that specifically present the neutralizing epitopes of the MPER and NHR region of gp41. These gp41 mimetics will be used to immunize rabbits, and the serum neutralizing Ab titers measured to determine the best MPER and NHR lead immunogens (Aim 1). In order to gain more specific knowledge about the immunogenicity of the neutralizing epitopes, rabbit mAbs that were elicited to these epitopes will be rescued by phage display and tested individually for neutralizing activity (Aim 2). The mAb panels will be used to probe the gp41 mimetics, and then modifications will be introduced so as to favorably display the neutralizing epitopes and occlude the non-neutralizing ones (Aim 3). In addition, in a collaborative effort, structures of the mAb Z13e1 and two HIV-1 neutralizing mAbs against the NHR region (rabbit and human) will be determined (Aim 4). In these ways, antibody access to the MPER and NHR region of gp41 will be precisely evaluated, which in turn should lead to improved gp41-based immunogens that will elicit more potent neutralizing Abs against HIV-1.